Retinitis pigmentosa is an untreatable, inherited retinal disease that leads to blindness. The disease initiates with the loss of night vision due to rod photoreceptor degeneration, followed by irreversible progressive loss of cone photoreceptors. Although loss of rods has substantial functional consequences such as night blindness and some constriction of visual fields, rod death by itself does not cause severe vision loss. Rather, it is the delayed wave of non-autonomous cone photoreceptor degeneration that results in blindness.
It has previously been shown that factors secreted from rods are essential for cone viability. One such trophic factor, RdCVF, has been identified (see U.S. Pub. No. 2004/0204350, Chalmel et al., BMC Molecular Biology 2007, 8:74; Leveillard et al., 2004, Nature Genetics, 36: 755-759; and Leveillard et al. at http://www.hal.inserm.fr/docs/00/31/23/78/DOC/Manuscript_on_April_15.doc). The feasibility of delivering RdCVF to the retina of a mouse model of retinitis pigmentosa (rd1) using recombinant AAV vectors has been evaluated. In vivo results suggests that efficient delivery of vector derived RdCVF protein to target photoreceptor cells requires novel secreted versions of the protein by the addition of specific signal peptides. Therapeutic applications of RdCVF have been explored, e.g., see WO 03/066810A2, EP 2027889A1, WO 2005/113586A2, Purification of secreted RdCVF and subsequent biochemical analysis confirms that RdCVF(LF) has anti-oxidative properties.
Recombinant expression of RdCVF at a large scale has been challenging. Effective expression of RdCVF from a gene therapy vector has also been challenging. Without being limited as to theory, it is theorized that post-translational events are largely responsible for the less than optimal levels of protein expression in either recombinant protein expression systems or in expression from a gene therapy vector. Following translation, proteins are routed specifically to secretory or endocytic pathways in the endoplasmic reticulum and traverse the Golgi network. Secreted proteins comprise a signal peptide, which is recognized during translation by the signal recognition pathway. Subsequently, the signal peptide is cleaved by a membrane—bound signal peptidase and the mature secreted protein enters a secretory vesicle. Thus secreted proteins are destined for the secretory pathway by recognition of their N-terminal signal peptides. The presently disclosed invention seeks to improve RdCVF expression profiles, including improving RdCVF secretion from cells both in vitro and in vivo.